We propose to use safe live-attenuated recombinant rabies vaccines to specifically target the induction of long-lasting immunity after administration of a single vaccine dose and extend the utility of the vaccines into the period when the virus has reached the CNS, clinical rabies is developing, and conventional post-exposure prophylaxis (PEP) with inactivated vaccines is no longer effective. Because live-attenuated rabies virus (RV) strains induce strong innate and adaptive immune responses that are capable of clearing the infection from the CNS, they will likely be effective for late stage rabies PEP. To overcome concerns regarding the safety of live- attenuated RV vaccines, we developed the triple RV G variant SPBAANGAS-GAS-GAS which is further attenuated, as illustrated by its apathogenicity for very young and immunoincompetent mice. Immunization with this variant induces strong innate and adaptive immune responses and is capable of preventing a lethal rabies encephalomyelitis in mice even when administered several days after infection with a highly pathogenic RV strain. Its high level of attenuation combined with its excellent post-exposure efficacy in mice makes SPBAANGAS-GAS-GAS a promising vaccine candidate not only for rabies PEP but also for the treatment of clinical rabies. To further examine the utility for single dose protective immunization and PEP, we will confirm the safety and stability of this highly attenuated RV variant and test its efficacy in hamsters and dogs, which have been demonstrated to be reliable animal models for pre- and post-exposure evaluation of rabies vaccines. In addition, we will elucidate the mechanisms that are responsible for the vaccine-mediated clearance of pathogenic RV from the CNS and further enhance the efficacy of live-attenuated RV vaccines to target RV-clearing immune effectors to the CNS by constructing recombinant RVs expressing molecules that enhance immune/inflammatory cell adhesion and extravasation across the neurovasculature/blood-brain barrier.